Interlaboratory comparison of autoradiographic DNA profiling measurements. 4. Protocol effects.

The observed total interlaboratory uncertainty in restriction fragment length polymorphism (RFLP) measurements is sufficiently small to be of little significance given current forensic needs. However, as the number of RFLP data increase, further reduction in the total uncertainty could help minimize the resources required to evaluate potential profile matches. The large number of data available enable quantitative estimation of the within-laboratory imprecision and among-laboratory bias contributions to the total uncertainty. Some small but consistent among-laboratory measurement biases can be attributed to specific procedural or materials differences. The bias direction is often fragment-specific and thus unpredictable for unknown samples. Actions that would minimize currently recognized sources of interlaboratory bias include the following: (1) all laboratories should use the same algorithm for data interpolation, (2) all laboratories should use the same sizing ladders, (3) each laboratory should prepare control DNA and sample DNA in the same manner and with the identical reagents, (4) all laboratories should adopt a uniform policy on ethidium bromide use, and (5) all laboratories should adopt the same control DNA sizing acceptability criteria.

Interlaboratory comparison of autoradiographic DNA profiling measurements. 2. Measurement uncertainty and its propagation.

Identifying the intrinsic sources of measurement uncertainty greatly facilitates control and further optimization of a measurement system. We have developed a model which quantitatively describes the observed interlaboratory variability of autoradiographic DNA band sizing. The model focuses on optical imaging measurements of band position and the calibration techniques used to convert measured band position to reported band size. The imaging component of measurement variability is described as a 0.05-0.2% standard deviation in determining the relative location of sample and calibration bands on a given film image. While developed solely with optical imaging information, the model is consistent with interlaboratory band sizing measurement variability observed with pristine samples. This interlaboratory variability can be modeled as a 0.2-0.4% standard deviation in the relative positions of sample and calibration bands across different electrophoretic gels. Further band sizing protocol standardization among laboratories would thus be expected to achieve at best a 2-fold reduction in interlaboratory band sizing variability.

An enzyme-linked immunosorbent assay (ELISA) for trinitrotoluene (TNT) residue on hands.

An enzyme-linked immunosorbent assay (ELISA) developed for the detection of trinitrotoluene (TNT) in munitions wastewater has been adapted to the detection of TNT residue on hands following contact. Using the procedure developed, as little as 50 pg of TNT could be detected. Accounting for sample size and dilution, the 50 pg equates to 15 ng of TNT recovered from the hands. Following contact with TNT, amounts ranging from 53 ng to more than 1500 ng were recovered from hands. The monoclonal anti-TNT antibodies showed no cross-reactivity with several other explosives or common contaminants. These preliminary results indicate promise for the development of a simple-to-use, immunoassay-based field test kit for TNT and, ultimately, other explosives.

A modification of the microplate method for reverse ABO typing of bloodstains and additional validation studies.

The results of additional validation studies of a sensitive microplate hemagglutination assay for ABO reverse grouping of bloodstains are presented. The results of the validation study demonstrate the reliability of the microplate assay for use in routine serological casework. Based on these studies, the microplate assay has now replaced the Lattes crust test for ABO reverse grouping of bloodstains in the FBI Laboratory.

The detection of soluble ABH blood group substances in semen and saliva using monoclonal blood grouping reagents.

Using an enzyme-linked immunosorbent assay (ELISA), this study investigated the use of monoclonal antibodies for detecting secreted ABH blood group substances in semen and saliva. The results demonstrated that the behavior of some monoclonals were unpredictable and often failed to detect the corresponding antigen in a number of the specimens tested. The suitability of the monoclonal reagents for detecting soluble blood group antigens could not be predicted by their behavior with red cell antigens. Consequently, care must be taken in the selection of monoclonal reagents for use in the detection of secreted blood group antigens.

A microplate method for reverse ABO typing of bloodstains.

A sensitive and reliable hemagglutination assay, using V-bottom microplates, is described for the detection of the ABO blood group alloantibodies in bloodstained material. When used in conjunction with an absorption-elution procedure, the microplate assay resulted in a 300% increase in the number of conclusive grouping results when compared to the Lattes crust test. The use of the microplate reverse grouping assay permits 24 specimens to be assayed conveniently on a single plate and eliminates the tedious and time-consuming microscopic examination required for the Lattes crust test.

The determination of sex from forcibly removed hairs.

The determination of sex from forcibly removed hairs in forensic science laboratories has, in the past, been based almost entirely on the presence or absence of the Y chromosome in the cells of the hair root sheath. Since the human male genotype is XY and the female is XX, a technique was devised that permits root sheath cells to be stained sequentially for the Y and then the X chromosome using quinacrine mustard. Following staining, the Y and the X chromosome fluorescence were observed, at pH 5.5 and 3.0, respectively, by epifluorescence. The X and Y chromosome counts obtained for a single hair root specimen were reported as a Y-X (Y minus X) score. The results reported show that specimens from males gave positive Y-X score while specimens from females gave negative Y-X scores. Results of an age study and blind trials were also reported.

A capillary tube method for the Lewis typing of red blood cells.

A simple and inexpensive capillary tube procedure, which can be applied in the forensic science laboratory, is described for the detection of the Lewisa (Lea) and Lewisb (Leb) antigens on red blood cells. This procedure will permit approximately 4000 tests to be performed from a single 2-mL bottle of Lewis antiserum.